The long-term objective of this proposal is to improve the therapy of glioblastoma multiforme (GBM), the most common and fatal of human gliomas. GBM result from the step-wise accumulation of genetic alterations and often arise from low-grade gliomas and anaplastic astrocytomas (AA). We found that 4 alterations (telomerase and Ras activation and p53/pRb inactivation) in combination allowed normal human astrocytes to form AA. Additional Akt activation, however, allowed formation of 5-fold larger, necrotic GBM. These results suggest that Akt contributes to GBM formation, a finding consistent with Akt activation in greater than 80 percent of GBM. While Akt enhances growth and angiogenesis, these effects were not seen in vitro, nor were they noted in vivo until tumors reached a critical size, at which point the Akt-expressing cells rapidly expanded into modestly vascularized (yet still hypoxic) GBM while the non-Akt expressing cells slowly expanded into AA. The growth-enhancing properties of Akt therefore appear to be unmasked by an additional event which we believe to be hypoxia. We hypothesize that the stimulus for the formation of GBM is hypoxia, and that Akt activation uniquely allows cells to grow and survive under these conditions. We will test this hypothesis with the following specific Aims: 1) to determine if the point at which the growth of model AA and GBM diverge corresponds with the onset of hypoxia, 2) to determine if conditional activation of Akt drives proliferation/survival of hypoxic Ras tumors, and if conditional suppression of Akt inhibits proliferation/survival of hypoxic Ras+Akt tumors, 3) to determine if hypoxia selects for cells expressing high Akt levels in vivo, 4) to determine if hypoxic conditions alone allow differential proliferation/survival of Ras+ Akt versus Ras astrocytes, and 5) to determine at the molecular level if the effects of hypoxia on cell cycle regulation and survival are modulated by Akt. Defining Akt function may help identify targets useful in GBM therapy.